SUMMARY About 40,000 women die each year from breast cancer in the US due to development of chemoresistance. Although ER/PR-positive breast cancer patients are treated effectively with endocrine therapy, chemotherapy is still one of the most effective and widely used means of treating breast cancer The patients in the triple- negative breast cancer (TNBC) group are dependent on chemotherapy as the sole therapy and their prognosis is poor due to inherent resistance. Her2 positive breast cancer patients are commonly treated with Herceptin but only in combination with chemotherapy. In addition, a significant fraction of breast cancer patients with large locally advanced cancers receives neoadjuvant therapy. Thus, there is clear unmet need to overcome chemoresistance in breast cancer patients. It is known that most cancer cells primarily use glycolysis as their main source of energy (ATP). However, this paradigm is currently changing in light of new studies supporting critical roles for mitochondria respiration in promoting cancer metastasis and chemoresistance. Thus, mitochondria are now being incorporated as targets into cancer treatment strategies but the challenge is the development of effective modulators of mitochondrial respiration that have minimal toxicity. We have identified MCJ (DnaJC15) as a mitochondrial protein that acts as an endogenous brake of the mitochondria respiration. MCJ is the first identified endogenous negative regulator of Complex I of the electron transport chain (ETC). Loss of MCJ expression promotes Complex I activity and production of mitochondrial ATP without affecting glycolysis rate. We have shown that MCJ expression is lost or markedly reduced in drug- resistant breast cancer cell lines. Recently, we have shown that loss of MCJ causes chemoresistance in vivo and that low MCJ in breast tumors correlates with poor chemotherapy response. Together, these studies indicate that restoring MCJ function in those breast cancer cells that have lost or reduced MCJ could be a novel approach to overcome chemoresistance. We have developed MCJ mimetics and our preliminary studies support the utility of these compounds as potential therapeutic drugs to safely reverse chemoresistance in breast cancer. The goal of this exploratory/developmental R21 application is to demonstrate that MCJ mimetics reverse resistance to a variety of standard chemotherapeutic drugs for breast cancer in vitro and in vivo using different mouse models, and cause no toxicity. We will determine: 1) the efficacy of MCJ mimetics in overcoming chemoresistance in vitro (Aim 1); the efficacy of MCJ mimetics in overcoming chemoresistance in vivo in mouse models (Aim 2) .